Prenatal Spina Bifida Repair: Defendable Trespassing of MOMS Criteria Results in Commendable Personalized Medicine

Introduction We hypothesize that after publication of the quintessence of the MOMS-Trial, eligibility criteria for prenatal spina bifida repair may be modified if a tenable argumentation underlies this decision. Methods Our first 154 fetal surgery patients were analyzed with particular focus on how many, which, and why the original eligibility criteria, set forth by the MOMS Trial Protocol, were disobeyed, and what the eventually detectable, negative and positive, impact of these deviations on outcomes was. Results A total of 152 patients (2 missing consent) were included (100%). In 69 patients (45.4%), a total of 89 eligibility criteria were disobeyed. In 54 (35.6%) cases, maternal criteria were concerned: Gestational age at operation of >25+6 weeks in 17 (11.2%), uterine pathologies in 13 (8.6%) women, preoperative BMI≥35 kg/m2 in 12 (7.9%), previous hysterotomy in 7 (4.6,%), previous prematurity in 3 (2%), HIV/hepatitis B in 2 (1.3%), psychosocial issues in 2 (1.3%), and placenta praevia in 1 (0.7%). In 32 (21.1%) cases, fetal criteria were disobeyed: Fetal anomaly unrelated to spina bifida in 19 (12.5%), no/minimal evidence of hindbrain herniation in 13 (8.6%), and severe kyphosis in 2 (1.3%). We could not identify cases where non-observation of criteria led to clear-cut maternal and/or fetal disadvantages. Conclusion This study shows that MOMS-Trial eligibility criteria for prenatal spina bifida repair should be modified or even abandoned with adequate medical and ethical argumentation, and with written parental informed consent after non-directive, full disclosure counseling. This clear cut change of paradigm is a necessity as it leads towards personalized medicine allowing more fetuses to benefit from fetal surgery than would have benefitted with the former published MOMS criteria in place

The spike gene is a major determinant for the SARS-CoV-2 Omicron-BA.1 phenotype.

Variant of concern (VOC) Omicron-BA.1 has achieved global predominance in early 2022. Therefore, surveillance and comprehensive characterization of Omicron-BA.1 in advanced primary cell culture systems and animal models are urgently needed. Here, we characterize Omicron-BA.1 and recombinant Omicron-BA.1 spike gene mutants in comparison with VOC Delta in well-differentiated primary human nasal and bronchial epithelial cells in vitro, followed by in vivo fitness characterization in hamsters, ferrets and hACE2-expressing mice, and immunized hACE2-mice. We demonstrate a spike-mediated enhancement of early replication of Omicron-BA.1 in nasal epithelial cultures, but limited replication in bronchial epithelial cultures. In hamsters, Delta shows dominance over Omicron-BA.1, and in ferrets Omicron-BA.1 infection is abortive. In hACE2-knock-in mice, Delta and a Delta spike clone also show dominance over Omicron-BA.1 and an Omicron-BA.1 spike clone, respectively. Interestingly, in naïve K18-hACE2 mice, we observe Delta spike-mediated increased replication and pathogenicity and Omicron-BA.1 spike-mediated reduced replication and pathogenicity, suggesting that the spike gene is a major determinant of replication and pathogenicity. Finally, the Omicron-BA.1 spike clone is less well-controlled by mRNA-vaccination in K18-hACE2-mice and becomes more competitive compared to the progenitor and Delta spike clones, suggesting that spike gene-mediated immune evasion is another important factor that led to Omicron-BA.1 dominance

The spike gene is a major determinant for the SARS-CoV-2 Omicron-BA. 1 phenotype

Variant of concern (VOC) Omicron-BA.1 has achieved global predominance in early 2022. Therefore, surveillance and comprehensive characterization of Omicron-BA.1 in advanced primary cell culture systems and animal models are urgently needed. Here, we characterize Omicron-BA.1 and recombinant Omicron-BA.1 spike gene mutants in comparison with VOC Delta in well-differentiated primary human nasal and bronchial epithelial cells in vitro, followed by in vivo fitness characterization in hamsters, ferrets and hACE2-expressing mice, and immunized hACE2-mice. We demonstrate a spike-mediated enhancement of early replication of Omicron-BA.1 in nasal epithelial cultures, but limited replication in bronchial epithelial cultures. In hamsters, Delta shows dominance over Omicron-BA.1, and in ferrets Omicron-BA.1 infection is abortive. In hACE2-knock-in mice, Delta and a Delta spike clone also show dominance over Omicron-BA.1 and an Omicron-BA.1 spike clone, respectively. Interestingly, in naïve K18-hACE2 mice, we observe Delta spike-mediated increased replication and pathogenicity and Omicron-BA.1 spike-mediated reduced replication and pathogenicity, suggesting that the spike gene is a major determinant of replication and pathogenicity. Finally, the Omicron-BA.1 spike clone is less well-controlled by mRNA-vaccination in K18-hACE2-mice and becomes more competitive compared to the progenitor and Delta spike clones, suggesting that spike gene-mediated immune evasion is another important factor that led to Omicron-BA.1 dominance

Molecular Basis of Increased Serum Resistance among Pulmonary Isolates of Non-typeable Haemophilus influenzae

Non-typeable Haemophilus influenzae (NTHi), a common commensal of the human pharynx, is also an opportunistic pathogen if it becomes established in the lower respiratory tract (LRT). In comparison to colonizing isolates from the upper airway, LRT isolates, especially those associated with exacerbations of chronic obstructive pulmonary disease, have increased resistance to the complement- and antibody-dependent, bactericidal effect of serum. To define the molecular basis of this resistance, mutants constructed in a serum resistant strain using the mariner transposon were screened for loss of survival in normal human serum. The loci required for serum resistance contribute to the structure of the exposed surface of the bacterial outer membrane. These included loci involved in biosynthesis of the oligosaccharide component of lipooligosaccharide (LOS), and vacJ, which functions with an ABC transporter encoded by yrb genes in retrograde trafficking of phospholipids from the outer to inner leaflet of the cell envelope. Mutations in vacJ and yrb genes reduced the stability of the outer membrane and were associated with increased cell surface hyrophobicity and phospholipid content. Loss of serum resistance in vacJ and yrb mutants correlated with increased binding of natural immunoglobulin M in serum as well as anti-oligosaccharide mAbs. Expression of vacJ and the yrb genes was positively correlated with serum resistance among clinical isolates. Our findings suggest that NTHi adapts to inflammation encountered during infection of the LRT by modulation of its outer leaflet through increased expression of vacJ and yrb genes to minimize recognition by bactericidal anti-oligosaccharide antibodies

Consensus guidelines for the use and interpretation of angiogenesis assays

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference

Oxytocin effects on chemosensory function in a clinical setting—a preliminary study

Background: Despite the large body of literature on the effects of oxytocin via the exchange of social chemo-signals, no previous study tested the effects of oxytocin stimulation during parturition on the mother’s chemical senses. Therefore, in the present study, we investigated the effects of oxytocin, administered via intramyometrial and intravenous injection, on general odor processing andgustatory functioning in women shortly after giving birth via Caesarean section. Methods: General olfactory and gustatory sensitivity, and subjectively perceived taste intensities and hedonic ratings, next to self-reported mood and nausea were assessed 1 day before the Caesarean section in 21 women and again a few hours after the oxytocin stimulations. Results: We found no changes in general olfactory sensitivity, self-reported mood, or nausea between test sessions. However, following oxytocin, women rated the quality of sweet taste as significantly more positive and tended to exhibit higher gustatory sensitivity. Conclusions: Although this study was performed in a highly controlled clinical environment, we cannot rule out potential confounders related to parturition, and our interpretation on the specific effects of oxytocin therefore is limited. However, our findings agree with the literature reporting oxytocin effects on chemo-sensory functions. We speculate that an increased preference for sweet taste might be particularly relevant during the early post-partum and breastfeeding period, facilitating thenecessary nutrient supply to the newborn via the intake of calorie-rich carbohydrate food

Mechanisms of Checkpoint Kinase Rad53 Inactivation after a Double-Strand Break in Saccharomyces cerevisiae

ABSTRACT In Saccharomyces cerevisiae , double-strand breaks (DSBs) activate DNA checkpoint pathways that trigger several responses including a strong G 2 /M arrest. We have previously provided evidence that the phosphatases Ptc2 and Ptc3 of the protein phosphatase 2C type are required for DNA checkpoint inactivation after a DSB and probably dephosphorylate the checkpoint kinase Rad53. In this article we have investigated further the interactions between Ptc2 and Rad53. We showed that forkhead-associated domain 1 (FHA1) of Rad53 interacts with a specific threonine of Ptc2, T376, located outside its catalytic domain in a TXXD motif which constitutes an optimal FHA1 binding sequence in vitro. Mutating T376 abolishes Ptc2 interaction with the Rad53 FHA1 domain and results in adaptation and recovery defects following a DSB. We found that Ckb1 and Ckb2, the regulatory subunits of the protein kinase CK2, are necessary for the in vivo interaction between Ptc2 and the Rad53 FHA1 domain, that Ckb1 binds Ptc2 in vitro and that ckb1 Δ and ckb2 Δ mutants are defective in adaptation and recovery after a DSB. Our data thus strongly suggest that CK2 is the kinase responsible for the in vivo phosphorylation of Ptc2 T376

Histone deposition promotes recombination-dependent replication at arrested forks

International audienceReplication stress poses a serious threat to genome stability. Recombination-Dependent-Replication (RDR) promotes DNA synthesis resumption from arrested forks. Despite the identification of chromatin restoration pathways after DNA repair, crosstalk coupling RDR and chromatin assembly is largely unexplored. The fission yeast Chromatin Assembly Factor-1, CAF-1, is known to promote RDR. Here, we addressed the contribution of histone deposition to RDR. We expressed a mutated histone, H3-H113D, to genetically alter replication-dependent chromatin assembly by destabilizing (H3-H4)2 tetramer. We established that DNA synthesis-dependent histone deposition, by CAF-1 and Asf1, promotes RDR by preventing Rqh1-mediated disassembly of joint-molecules. The recombination factor Rad52 promotes CAF-1 binding to sites of recombination-dependent DNA synthesis, indicating that histone deposition occurs downstream Rad52. Histone deposition and Rqh1 activity act synergistically to promote cell resistance to camptothecin, a topoisomerase I inhibitor that induces replication stress. Moreover, histone deposition favors non conservative recombination events occurring spontaneously in the absence of Rqh1, indicating that the stabilization of joint-molecules by histone deposition also occurs independently of Rqh1 activity. These results indicate that histone deposition plays an active role in promoting RDR, a benefit counterbalanced by stabilizing at-risk joint-molecules for genome stability

The Chromatin Assembly Factor 1 Promotes Rad51-Dependent Template Switches at Replication Forks by Counteracting D-Loop Disassembly by the RecQ-Type Helicase Rqh1

<div><p>At blocked replication forks, homologous recombination mediates the nascent strands to switch template in order to ensure replication restart, but faulty template switches underlie genome rearrangements in cancer cells and genomic disorders. Recombination occurs within DNA packaged into chromatin that must first be relaxed and then restored when recombination is completed. The chromatin assembly factor 1, CAF-1, is a histone H3-H4 chaperone involved in DNA synthesis-coupled chromatin assembly during DNA replication and DNA repair. We reveal a novel chromatin factor-dependent step during replication-coupled DNA repair: Fission yeast CAF-1 promotes Rad51-dependent template switches at replication forks, independently of the postreplication repair pathway. We used a physical assay that allows the analysis of the individual steps of template switch, from the recruitment of recombination factors to the formation of joint molecules, combined with a quantitative measure of the resulting rearrangements. We reveal functional and physical interplays between CAF-1 and the RecQ-helicase Rqh1, the BLM homologue, mutations in which cause Bloom's syndrome, a human disease associating genome instability with cancer predisposition. We establish that CAF-1 promotes template switch by counteracting D-loop disassembly by Rqh1. Consequently, the likelihood of faulty template switches is controlled by antagonistic activities of CAF-1 and Rqh1 in the stability of the D-loop. D-loop stabilization requires the ability of CAF-1 to interact with PCNA and is thus linked to the DNA synthesis step. We propose that CAF-1 plays a regulatory role during template switch by assembling chromatin on the D-loop and thereby impacting the resolution of the D-loop.</p></div